OBJECTIVE: To examine the effects of microRNA-34b-5p (miR-34b-5p) on bleomycin-induced lung fibrosis in mice and the underlying mechanism.
MATERIALS AND METHODS: TIMP3-deficient (Timp3−/−) and wild-type mice were administered with bleomycin before to detect the miR-34b-5p expression using quantitative Real Time-Polymerase Chain Reaction (qRT-PCR). Luciferase reporter assay was used to identify the target of miR-34b-5p in human lung fibroblast MRC-5. MiR-34b-5p was then silenced in vivo before lung histologic analysis and evaluation of extracellular matrix (ECM) genes as well as myofibroblast marker. The protein and mRNA expression levels were detected by Western blot and qRT-PCR, respectively.
RESULTS: We found that miR-34b-5p was significantly increased in lung tissues from bleomycin-stimulated mice. TIMP3 was identified as a direct target of miR-34b-5p by using dual-luciferase reporter assay, and enhanced expression of miR-34b-5p led to a decrease in TIMP3 whereas miR-34b-5p knockdown was responsible for TIMP3 elevation in MRC-5 cells. MiR-34b-5p knockdown in vivo attenuated the bleomycin-induced pulmonary fibrosis in wild-type mice, displayed by a reduced expression of Col1A1, fibronectin (Fn), and α-SMA. Furthermore, histological examination of lung sections also verified a diminishing fibrotic phenotype caused by the miR-34b-5p knockdown. But in Timp3-/- mice, down-regulation of miR-34b-5p did not exert an effect on the severe fibrotic lung injury after bleomycin exposure.
CONCLUSIONS: MiR-34b-5p knockdown appears to enhance the resistance to bleomycin by regulating its target gene TIMP3 during the pathogenesis of lung fibrosis.
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R.-P. Hu, Y.-Y. Lu, X.-J. Zhang
MiR-34b-5p knockdown attenuates bleomycin-induced pulmonary fibrosis by targeting tissue inhibitor of metalloproteinase 3 (TIMP3)
Eur Rev Med Pharmacol Sci
Vol. 23 - N. 5